The development of novel anti-cancer agents since 2000 includes about 1200 projects from in vivo lead optimisation, through pre-clinical phases, to Phase III clinical trials (Expert Opinion Emerging Drugs 6, 2001). Of these 1200 projects, less than 15% are under Phase I to Phase III clinical trials. Cytotoxic and cytostatic drugs and signaling pathway inhibitors are the two largest groups under development. Together with anti-angiogenic compounds and biologicals, e.g., monoclonal antibodies, they represent about 70% of anti-cancer drugs assayed in clinical phases since 2000. Further 19% of anti-cancer drugs under clinical trials belong to hormone therapy, cell cycle inhibitors, chemoprotective and histone deacetylase inhibitors. Compared to the above, only 3 anti-migratory (anti-metastatic) compounds entered clinical trials, i.e., 2% of those under clinical trials in oncology.
Despite the availability of more efficacious cytotoxic and cytostatic drugs and monoclonal antibodies that target cancer cells, the treatment of patients in advanced and/or metastatic disease remains highly unsatisfactory. The use of inhibitors of signaling pathways is further complicated because such pathways are not all activated at the same time during progression of a cancer. Rather, particular inhibitors need to be applied only when the corresponding target is present in the tumor tissue. Therefore, individual patients need to undergo molecular profiling of their tumors before any treatment, which decreases the efficiency and cost-effectiveness of the treatment, and introduces a delay before effective treatment can be started.
In view of the above, there remains a need for novel therapeutic approaches to combat cancer. New cellular targets are needed, as well as therapeutic molecules which can efficiently impinge on these targets. The cellular targets may preferably be involved in different cancers and may play a role in various disease mechanisms, such as, e.g., in migration of cancer cells and metastasis.
Galectins, originally named as galactose-specific lectins, is a family of 15 members in mammals. Each member of this family is expressed in a restricted set of normal and neoplastic tissues and is associated with distinct biological functions (Danguy et al. 2002). Galectins form homodimers or oligomers that can readily bridge N- and O-glycans as well as glycolipids present on cell surfaces with similar glycans in the ECM. In addition, cross-linking of β-galactoside-containing plasma membrane glycoconjugates modulates cell signaling, adhesion and survival. Although most galectins have been described as extracellular actors, intracellular functions have also been described. They have been found to play a number of important roles in biological processes including cellular processes involving cancers. As such, the role of galectins is very strongly tied to cancer and other proliferative diseases.
More specifically, the inventors contemplate that galectin-1 expression or over-expression in tumors or in tissues surrounding the tumors can be considered as a sign of malignant progression of the tumor and is often associated with poor prognosis for patients, often related to the dissemination of tumor cells at distance (metastasis) or in the surrounding normal tissue and to tumor immune-escape.
The inventors find that galectin-1 expression or over-expression may play particularly important role in non-small-cell-lung cancer (NSCLC), non-Hodgkin's (NH) lymphoma, pancreatic cancer, head & neck cancer, melanoma and glioma (brain tumors). These six cancer types may account for about 19.8% or more of all cancers encountered in female patients and about 37.2% or more of all cancers encountered in male patients. Hence, the number of female and male patients benefiting from an efficient targeting of these cancer types may be about 1,261,000 new cancer patients per year.
For example, galectin-1 is over-expressed in pancreatic ductal adenocarcinomas (Grutzmann et al. 2004; Shen et al. 2004) as compared to normal tissue and pancreatitis, a fact that relates to the level of differentiation of tumor cells (Berberat et al. 2001). Pancreatic stellate cells play a key role in the development of pancreatic fibrosis, a pathological feature of chronic pancreatitis and pancreatic cancer. Fritzner et al. 2005, showed that activation of rat pancreatic stellate cells is associated with increased expression of galectin-1 that modulates pancreatic stellate cell functions.
Galectin-1 expression has been demonstrated in head and neck squamous cell carcinomas (HNSCC). It is expressed within the invasive compartment of tumors (Gillenwater 1996) in relation with aggressiveness (Choufani et al. 1999).
Patients with non-small-cell lung cancer (NSCLC) are often positive for galectin-1 expression, among which adenocarcinomas figure prominently. The galectin-1 expression tends to increase with the progression of the malignancy and is an unfavorable independent prognostic factor that may relate to the proliferative activity of tumor cells (Szoke T et al. 2005; Gabius et al. 2002).
Recombinant galectin-1 added extracellularly to melanoma cells induces a dose-dependent increase of cell adhesion on laminin or fibronectin (van den Brule et al. 1995) and cell aggregation though interaction with glycoprotein 90K/MAC-2BP (Tinari N et al. 2001). The immunomodulatory effects of galectin-1 and the correlation between galectin-1 expression in cancer cells and the aggressiveness of these tumors (Rabinovich et al. 2002) make the inventors hypothesize that tumor cells may impair T-cell effector functions through secretion of galectin-1 and that this mechanism may contribute in tilting the balance towards an immunosuppressive environment at the tumor site. Rubinstein et al. 2004 advocated a link between galectin-1-mediated immuno-regulation and its contribution to tumor-immune escape. Blockade of the inhibitory effects of galectin-1 within melanoma tissue resulted in reduced tumor mass and stimulated the generation of a tumor-specific T-cell response in vivo. This supports the idea that galectin-1 may contribute to immune privilege of tumor by modulating survival or polarization of effector T cells, and suggest a potential molecular target for manipulation of T-cell apoptosis with potential implication in the therapeutic of cancer.
While the vessel walls of normal lymphoid tissues do not express galectin-1, the blood vessel walls in lymphomas express galectin-1 in relation with vascular density (D'Haene et al. 2005). Sezary cells, the malignant T cells in cutaneous T cell lymphoma (Sezary syndrome or mycosis fungoides) resist a variety of apoptosis—inducing agents, including galectin-1 induced apoptosis because of the loss of CD7 expression and altered cellular glycosylation. Recent evidence also indicates that galectin-1 (dGal-1) can induce the exposure of phosphatidylserine (an early apoptotic marker involved in the phagocytosis of apoptotic cells) on the plasma membrane of the human T leukemia MOLT-4 cells as well as on promyelocytic cell line and activated neutrophils, but that this does not result in cell death but prepares cells for phagocytic removal.
Galectin-1 has been reported to be the most important member of the galectin family in physiological brain processes (Danguy et al. 2002, Camby et al. 2001, Zanetta, 1998). The present invention is at least partly based on the finding of a direct implication of galectin-1 in the development of malignancy of human gliomas.
In patients bearing human glial tumors, the levels and patterns of expression of galectin-1 correlate with the development of malignancy (Camby et al. 2001). In a recent survey of clinical samples of high-grade astrocytic tumors, it was noticed that a low level of expression of galectin-1 in human malignant gliomas was associated with unusually long survival of such malignant glioma patients (Camby et al. 2001). Conversely, elevated levels of galectin-1 expression have been observed for highly invasive tumoral astrocytes, both in human surgical samples and animal models (Camby et al. 2001).
While increasing levels in galectin-1 expression correlate with malignancy development in human gliomas, such development of malignancy in human gliomas is associated with a marked decrease in galectin-3 expression (Camby et al. 2001). These data indicate different roles for galectin-1 and galectin-3 in the development of glioma malignancy. Therefore, the use of an anti-galectin-3 strategy for therapeutic purpose to combat cancer in general cannot be extrapolated to glioma in particular.
A direct involvement of galectin-1 in the aggressive behavior of malignant gliomas has been reported (Camby et al. 2002; Rorive et al. 2001, Gunnersen et al. 2000, Yamaoka et al. 2000).
For instance, the applicant has shown that in vitro, the addition of galectin-1 into the culture medium of U87 human glioblastoma cells markedly increased their migration capabilities (Camby et al. 2002, Rorive et al. 2001). These effects were associated with actin cytoskeleton reorganization and with increased expression in the small GTPase, RhoA (Camby et al. 2002). Conversely, human U87 glioblastoma cells constitutively expressing reduced levels of galectin-1 (U87/G1−) by means of stable transfection of an expression vector for antisense mRNA of galectin-1 were engineered. In vivo, intracranial grafting of U87/G1− cells into nude mice led to much longer survival in comparison with mice grafted with control cells (Camby et al. 2002). In vitro, U87/G1− cells were much less motile than parental (wt) and mock-transfected cells (Camby et al. 2002). Long-term deficiency in galectin-1 expression in these cells did not modify cell growth properties but impaired cell adhesion and invasiveness in Boyden chambers, and decreased expression and secretion and activity of matrix metalloproteinase-2. Matrix metalloproteinases-2 exerts marked roles in the development of malignancy of human gliomas (Rao 2003). The decrease in the levels of expression and secretion of galectin-1 in tumor astrocytes decreases the levels of expression and secretion of MMP-2 in these tumor astrocytes, a feature that will in turn decrease the capacity of tumor astrocytes to invade the brain parenchyma (Camby et al., 2002).
In order to further orient the study of the molecular mechanisms whereby galectin-1 promotes adhesion, motility and invasion of tumor astrocytes, the effect of stable transfection with antisense galectin-1 vector to mock-transfected and wild-type cells was also compared by cDNA microarray analysis. The expression of 91 genes (among 631 genes potentially involved in cancer) was increased by at least 2-fold. Confirmation of increased protein level was provided by immunocytochemistry for p21waf/cip1, cullin-2, p53, α9β1 integrin, ADAM-15 and MAP-2. Major differences in the expression pattern of α9β1 integrin and ADAM-15 proteins were also observed.
The use of galectin inhibitors for treating cancer in general has been reported. For instance, the use of an anti-galectin-4 or anti-galectin-9 therapeutic approach to combat certain types of cancers has been suggested. However, the present invention does not target galectin-4 or galectin-9.
US 2003/0109464 describes methods for inhibiting the growth and/or metastasis of a breast tumor in a subject by administering a therapeutic compound that binds and/or inhibits the activity of GAL-1 or GAL-4. The therapeutic compounds are amino acids or polypeptides coupled to one or more sugars. In some cases, parts of the GAL-1 and GAL-1 proteins themselves (e.g. parts of the binding domains) are used as therapeutic compounds, in other cases, non-GAL proteins (e.g. glycoamines) are used as therapeutic agents. The described approach is directed to the treatment of breast cancer and is not suitable or effective for the treatment of the above cancers, e.g., glioma, non-Hodgkin's lymphomas, non-small-cell-lung cancers, head & neck cancers, melanomas and pancreas cancers which consist of different pathologies.
WO 2004/091634 describes methods and compositions for augmenting treatment of different types of cancers and other proliferative disorders by combining the administration of an agent that inhibits the anti-apoptotic activity of galectin-3 (e.g., a “galectin-3 inhibitor”) so as to potentiate the toxicity of a chemotherapeutic agent. However, galectin-3 does not play a role in the development of glioma (Camby et al., 2001a). The level of expression of galectin-3 dramatically decreases during the progression of the glioma disease and when tumor malignancy develops (Camby et al., 2001a). Therefore the use of an anti-galectin 3 strategy for therapeutic purpose to combat cancer in general cannot be extrapolated to glioma in particular. The above-described approaches, which are based on the anti-apoptotic activity of galectin-3, and the inhibition thereof, are therefore not effective in the treatment of glioma.
In view of the above, it is clear that there remains a need in the art for therapeutic approaches to combat cancer, in particular malignant gliomas, pancreatic cancer, head and neck cancer, melanoma, non-small-cell lung cancer and non-Hodgkin's lymphoma. It is therefore an object of the present invention to provide nucleic acid compounds, compositions and methods for the treatment of cancers, in particular cancers associated with galectin-1 expression or overexpression, and in particular, for the treatment of malignant glioma, pancreatic cancer, head and neck cancer, melanoma, non-small-cell lung cancer and non-Hodgkin's lymphoma, which overcome at least some of the drawbacks of currently applied compositions and methods.